The effects of DNA sequence on enzymatic function has been an area of extensive research. Recent studies reveal strong DNA sequence effects on the 3~-exonuclease activity of Rrp1. The exonuclease acts on pyrimidine rich regions much more slowly than on purine rich regions. This enzymatic property is conserved between Rrp1 and it~s homologue, E. coli exonuclease III, however, its biological importance is not yet clear. Using a genetic screen, we isolated mutants of Rrp1 with altered DNA repair capacity. These mutants identify amino acids that modify the nuclease function and identify a glutamic acid residue essential for all nuclease function. Recently published crystallographic data supports this enzymatic analysis with structural information and indicates a critical role in the active site for the same glutamic acid residue. In recent enzymatic studies of Rrp1, a 3-phosphodiesterase assay was developed with higher sensitivity and specificity than standard assay methods. Application of this assay using extracts from yeast allowed identification of a novel repair enzyme with properties similar to Drosophila Rrp1. Partial purification yields a protein fraction enriched in a Mg++ dependent 3-phosphodieserase with associated class II AP endonuclease and weak 3-exonuclease. Further characterization of this yeast protein is in progress.